Achieving efficient protein expression in Trichoderma reesei by using strong constitutive promoters

Microbial Cell Factories

Research

Junxin Li¹  Miao Xing¹  Shaowen Wang¹  Shaowen Yu¹  Juan Wang¹  Gang Liu¹ 

[1] College of Life Science, Shenzhen Key Laboratory of Microbial Genetic Engineering, Shenzhen University, 518060, Shenzhen, China;

关键词: Trichoderma Reesei;    Xylanase;    Pyruvate decarboxylase;    Enolase;    Quantitative real-time PCR;   

DOI  :  10.1186/1475-2859-11-84

 received in 2012-02-09, accepted in 2012-06-07,  发布年份 2012

来源: Springer

【 摘 要 】

BackgroundsThe fungus Trichoderma reesei is an important workhorse for expression of homologous or heterologous genes, and the inducible cbh1 promoter is generally used. However, constitutive expression is more preferable in some cases than inducible expression that leads to production of unwanted cellulase components. In this work, constitutive promoters of T. reesei were screened and successfully used for high level homologous expression of xylanase II.ResultsThe transcriptional profiles of 13 key genes that participate in glucose metabolism in T. reesei were analyzed by quantitative real-time reverse-transcription polymerase chain reaction (RT-qPCR). The results indicated that the mRNA levels of pdc (encoding pyruvate decarboxylase) and eno (encoding enolase) genes were much higher than other genes under high glucose conditions. Recombinant T. reesei strains that homologously expressed xylanase II were constructed by using the promoters of the pdc and eno genes, and they respectively produced 9266 IU/ml and 8866 IU/ml of xylanase activities in the cultivation supernatant in a medium with high glucose concentration. The productivities of xylanase II were 1.61 g/L (with the pdc promoter) and 1.52 g/L (with the eno promoter), approximately accounted for 83% and 82% of the total protein secreted by T. reesei, respectively.ConclusionsThis work demonstrates the screening of constitutive promoters by using RT-qPCR in T. reesei, and has obtained the highest expression of recombinant xylanase II to date by using these promoters.

【 授权许可】

Unknown   
© Li et al.; licensee BioMed Central Ltd. 2012. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.